DESCRIPTION: (Applicant's Abstract) Drug addiction is one of the major health concerns of the present time. Increasing number of younger children find themselves drawn into the "drug" lifestyle of such drugs as amphetamine (AMPH). This research project will focus on the mode of action of AMPH as it pertains to the field of neuropharmacology with an emphasis on the role of protein kinase C (PKC) in AMPH-mediated DA release. Once the mechanism of AMPH is understood, pharmacological means can be employed in treating drug addiction. This research project will begin by examining whether the dopamine transporter (DAT) and norepinephrine transporter (NET) are regulated through phosphorylation and if inhibition of PKC will affect the AMPH-mediated DA release. PKC activation by AMPH will be tested and specific PKC isozymes involved will be elucidated. In addition, it will be determined if DAT or NET are attached to the cytoskeletal structures and if this attachment is modified by PKC and/or by AMPH. Also it will be investigated whether cultured cells can become sensitized to AMPH and how the sensitized state affects the cytoskeletal and vesicular arrangement of various proteins with DAT. This research project will utilize techniques such as protein-protein crosslinking, enzyme activity assays with 32P, immunoblotting, molecular assays for actin F and G, immunoprecipitation, HPLC and [3H]DA uptake assays.